PROJECT SUMMARY. Maintenance of an intact intestinal barrier is critical to prevent microbial activation of mucosal immunity. ?? intraepithelial lymphocytes (IEL) migrate extensively within the epithelial compartment to serve as a first line of defense against luminal antigens and invasive enteric microbes. Although ?? IELs can induce epithelial proliferation, whether ?? IELs regulate cell shedding has yet to be investigated. Our preliminary data generated for this application demonstrate ?? IELs are required for pathological cell shedding in response to systemic lipopolysaccharide (LPS) exposure; however, the mechanisms regulating this process remain unclear. This proposal seeks to elucidate the role of ?? IEL migratory behavior in pathological cell shedding and begin to define the molecular mechanisms by which ?? IELs contribute to these shedding events. Specifically, we propose to investigate the spatiotemporal dynamics of ?? IEL interactions with LPS-induced shedding events, the role of ?? IEL motility in regulating shedding, and whether local ?? IEL secretion of granzyme into the extracellular matrix facilitates release of enterocytes from the basement membrane as a critical component of the shedding process. Using an integrated approach that combines cutting edge imaging technologies with cell biology and immunological techniques, these studies will be the first to identify a direct role for ?? IELs in the regulation of pathological cell shedding. These experiments will not only elucidate a novel functional role for ?? IELs in the intestinal mucosa, but also provide the basis to investigate the extent to which specific subsets of ?? IELs contribute to maintenance of the epithelial barrier as a potential area of therapeutic development to aid in mucosal healing.